scholarly journals The Selection and Use of Sorghum (Sorghum propinquum) Bacterial Artificial Chromosomes as Cytogenetic FISH Probes for Maize (Zea maysL.)

2011 ◽  
Vol 2011 ◽  
pp. 1-16 ◽  
Author(s):  
Debbie M. Figueroa ◽  
James D. Davis ◽  
Cornelia Strobel ◽  
Maria S. Conejo ◽  
Katherine D. Beckham ◽  
...  
Keyword(s):  
Bac Library ◽  
Chromosome 9 ◽  
Bacterial Artificial Chromosomes ◽  
Fish Mapping ◽  
Artificial Chromosomes ◽  
Physical Maps ◽  
Sorghum Propinquum ◽  
Marker Loci ◽  
Single Clone

The integration of genetic and physical maps of maize is progressing rapidly, but the cytogenetic maps lag behind, with the exception of the pachytene fluorescencein situhybridization (FISH) maps of maize chromosome 9. We sought to produce integrated FISH maps of other maize chromosomes using Core Bin Marker loci. Because these 1 Kb restriction fragment length polymorphism (RFLP) probes are below the FISH detection limit, we used BACs from sorghum, a small-genome relative of maize, as surrogate clones for FISH mapping. We sequenced 151 maize RFLP probes and comparedin silicoBAC selection methods to that of library filter hybridization and found the latter to be the best. BAC library screening, clone verification, and single-clone selection criteria are presented along with an example of transgenomic BAC FISH mapping. This strategy has been used to facilitate the integration of RFLP and FISH maps in other large-genome species.

scholarly journals Defining the Deletion Size in Williams-Beuren Syndrome by Fluorescent In Situ Hybridization with Bacterial Artificial Chromosomes

10.5772/33238 ◽  
2011 ◽  
Author(s):  
Audrey Basinko ◽  
Nathalie Douet-Guilbert ◽  
Severine Audebert-Bellanger ◽  
Philippe Parent ◽  
Clemence Chabay-Vichot ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Fluorescent In Situ Hybridization ◽  
Bacterial Artificial Chromosomes ◽  
Artificial Chromosomes ◽  
Deletion Size

FISH of a maize sh2-selected sorghum BAC to chromosomes of Sorghum bicolor

Genome ◽  
1997 ◽  
Vol 40 (4) ◽  
pp. 475-478 ◽  
Author(s):  
Martha I. Gómez ◽  
M. Nurul Islam-Faridi ◽  
Sung-Sick Woo ◽  
Don Czeschin Jr. ◽  
Michael S. Zwick ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Sorghum Bicolor ◽  
Fluorescence In Situ Hybridization ◽  
Artificial Chromosome ◽  
Single Copy ◽  
Bacterial Artificial Chromosomes ◽  
Extra Copy ◽  
Artificial Chromosomes ◽  
Sorghum Plant

Fluorescence in situ hybridization (FISH) of a 205 kb Sorghum bicolor bacterial artificial chromosome (BAC) containing a sequence complementary to maize sh2 cDNA produced a large pair of FISH signals at one end of a midsize metacentric chromosome of S. bicolor. Three pairs of signals were observed in metaphase spreads of chromosomes of a sorghum plant containing an extra copy of one arm of the sorghum chromosome arbitrarily designated with the letter D. Therefore, the sequence cloned in this BAC must reside in the arm of chromosome D represented by this monotelosome. This demonstrates a novel procedure for physically mapping cloned genes or other single-copy sequences by FISH, sh2 in this case, by using BACs containing their complementary sequences. The results reported herein suggest homology, at least in part, between one arm of chromosome D in sorghum and the long arm of chromosome 3 in maize.Key words: sorghum, maize, shrunken locus, physical mapping, fluorescence in situ hybridization, bacterial artificial chromosomes.

scholarly journals Molecular characterization of the duplicated meristem identity genes HvAP1a and HvAP1b in barley

Genome ◽  
2005 ◽  
Vol 48 (5) ◽  
pp. 905-912 ◽  
Author(s):  
Liuling Yan ◽  
Jarislav von Zitzewitz ◽  
Jeffrey S Skinner ◽  
Patrick M Hayes ◽  
Jorge Dubcovsky
Keyword(s):  
Structural Changes ◽  
Growth Habit ◽  
Physical Map ◽  
Single Copy ◽  
Bacterial Artificial Chromosomes ◽  
Vernalization Gene ◽  
Artificial Chromosomes ◽  
Physical Maps ◽  
Meristem Identity

The vernalization gene VRN-1 has been identified as a MADS-box transcription factor orthologous to the meristem identity gene APETALA1 (AP1). A single copy of this gene was found in diploid wheat, but 2 copies were reported in barley. In this study, we present a detailed characterization of these 2 copies to understand their respective roles in the vernalization response. We identified 2 groups of barley bacterial artificial chromosomes (BACs), each containing 1 AP1 copy designated hereafter as HvAP1a and HvAP1b. A physical map of the VRN-H1 region showed that the HvAP1a BACs were part of the VRN-H1 region but that the HvAP1b BACs were not. Numerous structural changes were observed between the barley and wheat VRN-1 physical maps. In a population segregating for VRN-H1, the HvAP1a gene cosegregated with growth habit, suggesting that HvAP1a is the barley vernalization gene VRN-H1. The other copy, HvAP1b, was mapped on the centromeric region of chromosome 1H, the chromosome where vernalization gene VRN-H3 was previously mapped. We developed a mapping population segregating for VRN-H3 and showed that 2 molecular makers flanking HvAP1b locus were not linked to growth habit. The HvAP1b copy has a complete deletion of the first 2 exons, suggesting that it is a truncated pseudogene and not a candidate for VRN-H3. In summary, this study contributed a detailed physical map of the barley VRN-H1 region, showed several structural differences with the orthologous wheat region, and clarified the identity of the barley VRN-H1 gene.Key words: barley, vernalization, Vrn-1, physical map.

Segmental duplications within the Glycine max genome revealed by fluorescence in situ hybridization of bacterial artificial chromosomes

Genome ◽  
2004 ◽  
Vol 47 (4) ◽  
pp. 764-768 ◽  
Author(s):  
Janice Pagel ◽  
Jason G Walling ◽  
Nevin D Young ◽  
Randy C Shoemaker ◽  
Scott A Jackson
Keyword(s):  
In Situ Hybridization ◽  
Glycine Max ◽  
Repetitive Sequences ◽  
Genome Structure ◽  
Soybean Genome ◽  
Structure Evolution ◽  
Segmental Duplications ◽  
Bacterial Artificial Chromosomes ◽  
Artificial Chromosomes

Soybean (Glycine max L. Merr.) is presumed to be an ancient polyploid based on chromosome number and multiple RFLP fragments in genetic mapping. Direct cytogenetic observation of duplicated regions within the soybean genome has not heretofore been reported. Employing flourescence in situ hybridization (FISH) of genetically anchored bacterial artificial chromosomes (BACs) in soybean, we were able to observe that the distal ends of molecular linkage group E had duplicated regions on linkage groups A2 and B2. Further, using fiber-FISH, it was possible to measure the molecular size and organization of one of the duplicated regions. As FISH did not require repetitive DNA for blocking fluorescence signals, we assume that the 200-kb genome region is relatively low in repetitive sequences. This observation, along with the observation that the BACs are located in distal euchromatin regions, has implications for genome structure/evolution and the approach used to sequence the soybean genome.Key words: soybean, genome evolution, FISH, chromosomes, physical mapping.

scholarly journals Cross-Species RNAi Rescue Platform in Drosophila melanogaster

Genetics ◽  
2009 ◽  
Vol 183 (3) ◽  
pp. 1165-1173 ◽  
Author(s):  
Shu Kondo ◽  
Matthew Booker ◽  
Norbert Perrimon
Keyword(s):  
Cell Culture ◽  
Drosophila Melanogaster ◽  
Large Scale ◽  
Transgenic Animals ◽  
Selection Marker ◽  
Bacterial Artificial Chromosomes ◽  
Loss Of Function ◽  
Artificial Chromosomes ◽  
Target Effects

RNAi-mediated gene knockdown in Drosophila melanogaster is a powerful method to analyze loss-of-function phenotypes both in cell culture and in vivo. However, it has also become clear that false positives caused by off-target effects are prevalent, requiring careful validation of RNAi-induced phenotypes. The most rigorous proof that an RNAi-induced phenotype is due to loss of its intended target is to rescue the phenotype by a transgene impervious to RNAi. For large-scale validations in the mouse and Caenorhabditis elegans, this has been accomplished by using bacterial artificial chromosomes (BACs) of related species. However, in Drosophila, this approach is not feasible because transformation of large BACs is inefficient. We have therefore developed a general RNAi rescue approach for Drosophila that employs Cre/loxP-mediated recombination to rapidly retrofit existing fosmid clones into rescue constructs. Retrofitted fosmid clones carry a selection marker and a phiC31 attB site, which facilitates the production of transgenic animals. Here, we describe our approach and demonstrate proof-of-principle experiments showing that D. pseudoobscura fosmids can successfully rescue RNAi-induced phenotypes in D. melanogaster, both in cell culture and in vivo. Altogether, the tools and method that we have developed provide a gold standard for validation of Drosophila RNAi experiments.

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